Multiple reaction monitoring (MRM) is a highly selective, high sensitivity mass spectrometry mode for detecting the presence of particular species in a mixture. In the pharmaceutical industry, many small molecule analytes (e.g. drug metabolites, hormones, and pesticides) are routinely measured in high throughput with MRM's great precision. MRM based assays are now being tested and developed for large biomolecules (peptides and proteins) in the field of proteomics. These advances are significant because MRM proteomics provides a high-throughput, flexible, and relatively inexpensive alternative to traditional protein assays. The use of MRM proteomics is particularly appealing compared to traditional protein anti-body based approaches because of the minimal time required to design and test an assay (fast instrument configuration) and the ability to multiplex thousands of assays. This proposal will develop a next-generation MRM software package that will provide full support during all aspects of MRM experiments, including transition selection, instrument optimization, data collection, and curation. The long-term goal is to develop a licensable, instrument independent software package for users with any level of expertise in targeted proteomics. Testing and development will be done through collaboration with research groups at the forefront of technology development in the field. The software suite will fill a void for proteomics laboratories globally as a comprehensive software package currently does not exist. The state of the art in MRM proteomics includes both focused academic applications as well as vendor-specific basic software packages bundled with instrument sales. However, the functionality provided by these tools is limited and few if any commercial quality tools are accessible to researchers looking to develop targeted proteomic efforts. The timing of this proposal is opportune to capitalize on the growing interest about MRM in the proteomics field where such an application will accelerate further use of MRM-based assays. This proposal's objective and implementation of the goals aligns with the NIH's mission in two key areas. First, the described MRM software would significantly accelerate development of MRM-based assays for protein level biomarkers and take full advantage of current instrumentation. The MRM approach is an established, well received technique and the discipline of proteomics is just starting to utilize the practice. The most recent ASMS conference included dozens of posters and talks describing MRM developments in the proteomics field. In addition, there are currently over 20 funded NIH grants supporting MRM or targeted proteomic efforts listed in the CRISP database. Second, the proposal will directly aid at least two research groups at leading institutions that are using the MRM methodology in proteomic analyses of human diseases. These collaborations will accelerate the respective groups'advances in early detection of heart disease and various human cancers. PUBLIC HEALTH RELEVANCE: Protein identification and abundance measurement enables researchers and medical personnel to better diagnose and understand medical conditions. This proposal develops software that finds the subtle changes in proteins that may cause disease.